During the past year, the Retroviral Diseases Section has conducted research on AIDS-related malignancies and also on HIV disease. Much of the work on AIDS-related malignancies has focused on tumors associated with Kaposis sarcoma-associated herpesvirus (KSHV), also called human herpesvirus-8 (HHV-8). This virus is the cause of Kaposis sarcoma (KS), primary effusion lymphoma (PEL) and multicentric Castlemans disease (MCD). We have found that hypoxia can activate latent KSHV to undergo lytic replication and that several genes of KSHV are specifically upregulated by hypoxia. We had previously found that a gene of unknown function, ORF34, is specifically activated by hypoxia. ORF34 is part of a cluster of genes (ORF34 to 37). We have dissected the molecular structure of this gene cluster and its upregulation by hypoxia. One of these genes, ORF36, can phosphorylate ganciclovir, and activation of this gene by hypoxia may be able to be used to therapeutic benefit in KSHV-associated tumors, especially PEL. In addition, we have found that a KSHV-encoded thymidine kinase (ORF21) is upregulated inKSHV-infected cells exposed to hypoxia. This gene can phosphorylate zidovudine (AZT) to a form that is toxic for cells. We have further found that phosphorylation of these drugs is increased in PEL cells exposed to hypoxia and that clinically attainable concentrations of these drugs can kill the PEL cells. This may provide a novel targeted strategy to treat PEL (which arises in hypoxic environments) and MCD (in which lytic KSHV genes are activated). We have also found that certain drugs used to treat Kaposis sarcoma (KS), such as doxorubicin, can activate the virus. We are also conducting several clinical trials to bring these discoveries into the clinic and explore other novel approaches to HIV-associated tumors. We have previously found that the cytokine IL-12 has long-acting activity in KS have also conducted a trial to study the combination of IL-12 and a liposomal anthracycline in patients with advanced KS. We are also studying antibody to VEGF (bevacizumab) as a therapeutic agent in KS as wel as BAY 43-9007 (sorafineb) . We have initiated trials to study the natural history of multicentric Castleman's disease (MCD), which is also caused by KSHV, as well as the use of AZT and valganciclovir to treat this disease. We are also studying infusional chemotherapy as therapy for AIDS-associated lymphoma in collaboration with the Metabolism Branch, and have initiated a trial of methobased chemotherapy for AIDS-associated lymphoma. With regard to HIV, we have been focusing on two areas: the HIV protease and developing an immune response to peptide sequences that confer resistance to anti-HIV drugs. Our group previously found that glutathiolation of a conserved cysteine at the HIV protease dimmer interface (Cys 95) abolishes HIV-1 protease activity. HIV virions with mutations of Cys 95 have been generated and we are studying the effects that these mutations have on the fitness of HIV under varying conditions. This work on HIV protease has identified the dimer interface as a potential therapeutic target, and we have focused on the dimer interface as a novel target. We showed that a peptide could be designed that interfered with HIV protease dimerization and that this peptide could block HIV viral production from infected cells. We are exploring the effect of these inhibitors on the Gag-Pol polyprotein, which needs to form a dimer and self-cleave itself to form active protease. We have also been exploring the possibility of designing a peptide vaccine to HIV that could induce an immune response to a viral sequence that confers resistance to an HIV drug. In collaboration with Dr. Jay Berzofsky, we have engineered such an peptide that can induce a response against the M184V sequence of HIV reverse transcriptase that confers resistance to lamivudine, and have initiated a clinical trial to explore this as a therapeutic vaccine.